Myocardial ischemia is a serious pathological condition of the heart muscle resulting from a lack of adequate arterial blood flow. The central events in ischemia are failure of the energy-producing systems, and metabolite accumulation due to lack of flow, followed by a cascade of interrelated secondary events leading to irreversible damage and necrosis. Brief periods of ischemia lead to a depression of heart functions and damage which is totally or partially reversible. I will examine both the irreversible and reversible forms of ischemic damage in two experimental models - the isolated, isovolumic, working rabbit heart and cardiac skinned muscle fiber bundles, to test the following hypotheses: 1) In severe ischemia, there is damage to the actomyosin complex, the calcium ion control myofilament proteins, or the sarcoplasmic reticulum (SR). 2) In mild ischemia, there is little damage to these structures, and the decrease in cardiac function ("stunned myocardium") is due to other anormalities such as sarcolemmal damage or inappropriate intracellular milieu. 3) Reperfusion may initially exacerbate this damage and then partially or fully reverse it. Similar hypotheses will be tested in hypoxic and reoxygenated models. I will use cardiac skinned fiber bundles (no functional sarcolemma) to identify sites of subcellular damage in ischemic, reperfused, hypoxic and reoxygenated hearts. Serial sample biopsies will be taken from an isolated heart, cardiac skinned fibers prepared, and tested for a number of contractile functions. In the ischemic model, for example, biopsies will be taken during the well-oxygenated control period, at increasing times of normothermic global ischemia, and during the period of reperfusion. I will correlate the decline in contractile functions in the heart such as developed pressure, dP/dt and diastolic stiffness with the decline in contractile functions in cardiac skinned fiber bundles such as maximum isometric tension, dT/dt, stiffness, calcium sensitivity of tension and the release of Ca2+ from fiber SR. Skinned fiber preparations from control, ischemic, and reperfused hearts will be tested for various contractile functions both in standard physiological ionic conditions, and in other solutions more nearly approximating what are thought to be the specific ionic conditions in ischemic and reperfused cardiac cells (low MgATP, low pH, abnormally high intracellular Ca2+). This work should lead to a better definition of the sites of ischemic and hypoxic damage in cardiac cells, and of a better estimation of the relative roles of ischemic and reperfusion damage in ischemic heart disease.